Is it possible to use the mouse CNS classifier as a starting point for human or rat CNS tissue?

[James Hong]

Hey everyone,

Trying to use the mouse classifier on Garnett for our rat / human tissue data:

colData(wt_rn)$garnett_cluster <- clusters(wt_rn)

mouse_wt_rn_classifier <- train_cell_classifier(cds=wt_rn,marker_file =
                                                  "./mmBrain_markers.txt",
                                                db=org.Rn.eg.db::org.Rn.eg.db,
                                                cds_gene_id_type = "ENSEMBL",
                                                num_unknown = 50,
                                                marker_file_gene_id_type = "SYMBOL",
                                                cores=4)

For some reason, when I remove cores=4 (I've run my own training classifier on cores=4 without fail), it classifies, but when I add it it throws:

<simpleError in file(con, "w"): all connections are in use>
GLMNET failed

---

When removing cores = 4, it seems to classify:

training_sample
              CNS glia           Immune cells Neural crest-like glia                Neurons                Unknown         Vascular cells
                   500                    500                    500                    500                     20                    279
Model training finished.
training_sample
                               Cerebellum neurons Cholinergic monoaminergic and peptidergic neurons
                                               26                                                18
                    Di- and mesencephalon neurons                                   Enteric neurons
                                              111                                               500
                                Hindbrain neurons                                   Immature neural
                                               65                                                14
                       Peripheral sensory neurons                               Spinal cord neurons
                                               51                                                11
                       Telencephalon interneurons                  Telencephalon projecting neurons
                                              114                                               112
                                          Unknown
                                               20
Model training finished.
training_sample
               Microglia Perivascular macrophages                  Unknown
                     500                      500                       20
Model training finished.
training_sample
                    Pericytes                       Unknown Vascular leptomeningeal cells  Vascular smooth muscle cells
                          134                            20                            40                           228
Model training finished.
training_sample
Astroependymal cells     Oligodendrocytes              Unknown
                 236                  500                   20
Model training finished.
training_sample
                    Enteric glia  Enteric mesothelial fibroblasts      Olfactory ensheathing cells Oligodendrocytes precursor cells
                             500                                2                              500                               19
                  Satellite glia                          Unknown
                             418                               20
The following cell types do not have enough training cells and will be dropped:  Enteric mesothelial fibroblasts
Model training finished.
training_sample
 Granular layer interneurons cerebellum Molecular layer interneurons cerebellum                  Neuroblasts cerebellum
                                      4                                     104                                       8
                                Unknown
                                     20
The following cell types do not have enough training cells and will be dropped:  Granular layer interneurons cerebellum
Model training finished.
training_sample
Cholinergic and monoaminergic neurons                   Peptidergic neurons                               Unknown
                                  500                                    44                                    20
Model training finished.
training_sample
Di- and mesencephalon excitatory neurons Di- and mesencephalon inhibitory neurons                                  Unknown
                                     367                                       90                                       20
Model training finished.
training_sample
Excitatory neurons hindbrain hind    Inhibitory neurons spinal cord                           Unknown
                               69                               500                                20
Model training finished.
training_sample
Spinal cord excitatory neurons                        Unknown
                           164                             20
Model training finished.
training_sample
Non-glutamatergic neuroblasts                       Unknown
                           95                            20
Model training finished.
training_sample
         Olfactory inhibitory neurons Telencephalon inhibitory interneurons                               Unknown
                                  342                                   500                                    20
Model training finished.
training_sample
              Dentate gyrus granule neurons Telencephalon projecting excitatory neurons Telencephalon projecting inhibitory neurons
                                         41                                         204                                         500
                                    Unknown
                                         20
Model training finished.
training_sample
Cholinergic enteric neurons   Nitrergic enteric neurons                     Unknown
                        500                           2                          20
The following cell types do not have enough training cells and will be dropped:  Nitrergic enteric neurons
Model training finished.
training_sample
  Peripheral sensory neurofilament neurons Peripheral sensory non-peptidergic neurons     Peripheral sensory peptidergic neurons
                                       315                                         19                                        500
                                   Unknown
                                        20
Model training finished.
No cells classified as Sympathetic neurons. No subclassification
training_sample
                           Astrocytes              Choroid epithelial cells                       Ependymal cells
                                  500                                   500                                   348
Subcommissural organ hypendymal cells                               Unknown
                                    2                                    20
The following cell types do not have enough training cells and will be dropped:  Subcommissural organ hypendymal cells
Model training finished.
training_sample
Committed oligodendrocytes cells          Mature oligodendrocytes  Myelin forming oligodendrocytes    Newly formed oligodendrocytes
                             180                              500                              115                              500
                         Unknown
                              20
Model training finished.
training_sample
                         Adrenergic cell groups of the medulla                       Afferent nuclei of cranial nerves VI-XII
                                                           216                                                             14
Cholinergic neurons septal nucleus Meissnert and diagonal band                       Dopaminergic neurons periaqueductal grey
                                                           501                                                             47
         Glutamatergic projection neurons of the raphe nucleus                           Noradrenergic neurons of the medulla
                                                            11                                                             24
                         Orexin-producing neurons hypothalamus                                      Pmch neurons hypothalamus
                                                             3                                                              4
                                          Serotonergic neurons                                                        Unknown
                                                           500                                                             20
The following cell types do not have enough training cells and will be dropped:  Orexin-producing neurons hypothalamus Pmch neurons hypothalamus
Model training finished.
training_sample
    Central canal neurons spinal cord     Inhibitory neurons septal nucleus      Inhibitory neurons telencephalon
                                    6                                    31                                   120
Oxytocin-producing cells hypothalamus      Peptidergic neurons hypothalamus                               Unknown
                                    6                                    36                                    20
The following cell types do not have enough training cells and will be dropped:  Central canal neurons spinal cord Oxytocin-producing cells hypothalamus
Model training finished.
training_sample
   Excitatory neurons hindbrain Excitatory neurons hypothalamus     Excitatory neurons midbrain     Excitatory neurons thalamus
                             44                             500                              50                              33
                        Unknown
                             20
Model training finished.
training_sample
   Inhibitory neurons hindbrain di Inhibitory neurons hypothalamus di     Inhibitory neurons midbrain di                            Unknown
                                 6                                126                                500                                 20

---

After it does the training, when going to classify cells:

wt_rn <- classify_cells(wt_rn, mouse_classifier,
                           db = org.Rn.eg.db,
                           cluster_extend = TRUE,
                           cds_gene_id_type = "ENSEMBL")

All of the cells are marked as "Unknown" rather than the cells it seems to have classified using the mouse marker files from Garnett.

Is it just there is no way to adapt the classifiers across-species? Or am I missing something?

Thanks!

James

[Hannah Pliner]

Hi James,

Can you clarify what gene names you're using for this cross-species classification? i.e. are the gene names in wt_rn ensembl rat IDs? By default, the classifier will be generated using ensembl IDs in whatever species is provided in the db.

Hannah

[James Hong]

Hey Hannah,

My intention was hoping to use the mouse CNS marker file (gene symbols) alongside the wt_rn ensembl IDs (translated using the rnor ensembl database to gene symbols) to produce a classifier that is capable of deciphering the rat data we have.

Is there a way to make this happen? I tried to classify cells directly from the .RDS but as expected as the mouse ensembl's didn't exist in the wt_rn cds it failed.

Thank you,

James

[Hannah Pliner]

Hi James,

If the gene names are compatible (and mark the same thing in rat as in mouse) then I don't see why there would be a problem. Perhaps try classifying using just a few cell types and known markers to see if the issue is with the mouse versus rat markers or something else? Do all the gene names get translated? i.e. when you run check_markers, does it look like all the marker genes are present in the training CDS?

Hannah